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Study on the Effects of Generated Trans-arachidonic Acids on Retinal Capillary during Nitrative Stress in Diabetic Rats and the Mechanisms

Author: XuLin
Tutor: XuXun
School: Shanghai Jiaotong University
Course: Ophthalmology
Keywords: Diabetic retinopathy Trans-arachidonic acid Nitrative stress Reactive Nitrogen Species NO2· Thrombospondin-1 Retinal capillary Apoptosis Cell culture GC/MS
CLC: R774.1
Type: PhD thesis
Year: 2008
Downloads: 141
Quote: 0
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Abstract


Purpose: To explore the role of nitrative stress on the pathogenesis of diabetic retinopathy. To investigate whether the TAAs change in diabetic conditions and effects to capillary cells and its possible mechanisms.Method: (1) Diabetes was induced by intraperitoneal injection of Streptozoticin (STZ) at 60 mg/kg body weight. The amount of TAAs and AA in serum of diabetic rats was measured by gas chromatography and mass spectrometry (GC/MS) method and the ratio of peak area of TAAs to AA with selected ion of 79 was estimated by group t-test. (2) Bovine retinal pericytes were cultured. Pericytes were treated 24h with different 14E-AA (5μM or 10μM) concentration. Apoptosis was measured by TUNEL, MTT and flow cytometry. (3) Expression of TSP-1 in retina was examined by immunohistochemical method and Western blot. Bovine retinal endothelial cells were cultured with selective culture media. At different time points after treated with high glucose, expresssion of TSP-1 and phosphorylation of ERK 1/2 of endothelial cells was examined by Western blot. The mRNA expression of TSP-1 was determined by real-time RT-PCR. Result: (1) The TAAs/AA was significantly increased in serum at the time point of 8w, 12w and 16w after induced by STZ (P <0.05). (2) TAAs couldn’t induce the apoptosis of retinal pericytes (P >0.05). (3) Expression of TSP-1 in retina of diabetic rats was elevated with longer duration of diabetes (checked the time point of 2w, 4w, 8w, 12w and 16w-diabetic rats). The expression of TSP-1 tended to physiologically age-related elevated, but less significant than diabetic group. A increase of TSP-1 expression in endothelial cells was observed after high glucose treated 48h. After high glucose treated, ERK 1/2 was activated since 30min. mRNA of TSP-1 was elevated after high glucose treated 48h than control, but decreased at 5d (P <0.05).Conclusion: (1) High glucose could induce nitrative stress. Function as specific lipid markers or mediators of the damage to biological membrane by NO2, TAAs generated in the serum of diabetic rats and a possible mechanism of microvascular injury exits in diabetic conditions. Thus, intervention targeting the formation of TAAs or NO2·may have potential therapeutic applications in conditions such as ischemic retinopathy and other microangiopathies. (2) Pericytes were unaffected by exposure to TAAs. (3) Under high glucose condition, TAAs induced microvascular damage through up-regulation of TSP-1, ERK1/2 seems to be required for TAAs signaling.

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CLC: > Medicine, health > Ophthalmology > Retina and optic nerve diseases > Retinal disease
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